This invention relates to high-density arrays on polymeric substrates and, more particularly to high-density arrays including mask layers and methods of manufacturing the same.
Arrays may be used in a variety of applications, such as gene sequencing, monitoring gene expression, gene mapping, bacterial identification, drug discovery, and combinatorial chemistry. Many of these applications involve expensive and oftentimes difficult to obtain samples and reagents. Accordingly, high density, miniaturized arrays are desirable because the use of such arrays may dramatically increase efficiency with respect to limited or expensive samples when compared to standard arrays, such as a 96 well plate. For example, a 96 well plate may require several hundred microliters of sample per well to run a diagnostic experiment whereas a miniaturized array would require only a fraction of that sample for the entire array. In addition to the reduction of volume, miniaturization allows hundreds or thousands of tests to be performed simultaneously. Furthermore, a high-density array may be more versatile than a standard array because of the wide variation of chemistries that may be present on a single array.
The arrays are typically used to determine the presence of a desired analyte by detecting an electromagnetic signal, e.g., fluorescence, emitted by the analyte in response to excitation. The excitation may be provided in the form of electromagnetic energy from, e.g., a laser. One potential problem with the arrays is that the electromagnetic energy used to excite the analyte may also excite other materials present in the substrate or other portions of the array, causing them to emit an electromagnetic signal similar to that emitted by the desired analyte. For example, where the array is formed on a polymeric substrate that fluoresces in the same or similar wavelength regions as the analyte, a relatively high background electromagnetic signal can be produced by the substrate that reduces the signal-to-background ratio. A lower signal-to-background ratio can make accurate detection of the desired analyte more difficult.
Another potential problem with the arrays is that the substrate may scatter light. For example, nylon and nitrocellulose membranes commonly used for many arrays are white and suffer tremendously from light scatter when used with optical scanners used for, e.g., fluorescence intensity measurements.
Although glass substrates exhibit low fluorescence and light scatter, glass cannot be used to manufacture arrays in mass production processes (such as continuous web manufacturing) because of its brittleness and low ductility.
The present invention provides, in some aspects, high-density arrays including mask layers and methods of manufacturing the same. The arrays are formed on a polymeric substrate and include a mask layer. Various linking agents, linking agent coatings, and/or reactants may be provided on the mask layer. Where present, the linking agents and reactants preferably operate together to capture a desired analyte which can then be detected based on an electromagnetic signal, e.g., fluorescence, that is emitted by the analyte in response to excitation energy incident on the array.
In some instances, the mask layer preferably reduces or prevents transmission of the excitation energy through the mask layer to the underlying substrate. In other instances, the mask layer may be used to reduce or prevent the transmission of electromagnetic energy from beneath the analyte, e.g., the substrate, that is similar to the electromagnetic signal emitted by the desired analyte in response to the excitation energy. In either case, with the mask layer in place, the electromagnetic signals emitted from the surface of the array can generally be attributed to excitation of the analyte captured on the array, rather than the underlying substrate or other portions of the array.
The substrates and mask layers used in connection with the high-density arrays also provide for reduction in the surface area of the arrays during manufacturing. The reduced surface area can provide a number of advantages including increases in the binding site density on the array, increases in the optical density of the mask layer (to improve the masking capabilities discussed above), and an undulated surface on the array. The undulated surface can improve reactant density on the surface of the arrays coating, thereby increasing the density of the captured analyte. As a result, accurate detection of the desired analyte captured on the arrays can be significantly improved. It is believed that, in some cases, the signal-to-background ratio obtained using high-density arrays of the present invention can be significantly enhanced, e.g., the signal-to-background ratio can exceed 40 or more.
In one aspect, the present invention provides an article for use in manufacturing an array, the article including a polymeric substrate; a mask layer on the article having a projected surface area and a topographical surface area that is greater than the projected surface area; and linking agents on the mask layer.
In another aspect, the present invention provides an array including a polymeric substrate; a mask layer on the array having a projected surface area and a topographical surface area that is greater than the projected surface area; linking agents on the mask layer; and reactants affixed to the linking agents to form binding sites on the array.
In another aspect, the present invention provides an article including a polymeric substrate and a layer on the article having at least one metal or metallic compound, the layer located on a first major surface of the substrate, wherein the layer has a projected surface area and a topographical surface area that is greater than the projected surface area.
In another aspect, the present invention provides a method of forming an article by providing a polymeric substrate having a first major surface with a first surface area; providing a layer on the first major surface of the substrate, wherein the layer includes at least one metal or metallic compound, and further wherein the layer has a projected surface area and a topographical surface area that are equivalent; and relaxing the substrate to reduce the first surface area after providing the layer thereon, wherein, after relaxing, the topographical surface area of the layer is increased such that it is greater than the projected surface area of the layer.
In another aspect, the present invention provides an article for use in manufacturing an array, the article including a polymeric substrate and a mask layer having ink and linking agents located on the article, wherein the mask layer has a projected surface area and a topographical surface area that is greater than the projected surface area.
In another aspect, the present invention provides an array including a polymeric substrate, a mask layer having ink and linking agents on the array, wherein the mask layer has a projected surface area and a topographical surface area that is greater than the projected surface area; and reactants affixed to the linking agents to form binding sites on the array.
In another aspect, the present invention provides a method of manufacturing an array by providing a polymeric substrate having a first major surface with a first surface area; providing a mask layer on the first major surface of the substrate, the mask layer exhibiting an original optical density for light of selected wavelengths; providing binding sites on the mask layer; and relaxing the substrate to reduce the first surface area after providing the mask layer, wherein the relaxing increases the optical density of the mask layer to a relaxed optical density that is greater than the original optical density for light of selected wavelengths.
These and other features and advantages of the invention are described in more detail below with respect to illustrative embodiments of the invention.